PROJECT SUMMARY Liver kinase B1 (LKB1) is a key regulator of cellular energy homeostasis and well known tumor suppressor gene in many cancers, particularly lung cancer, where it is deleted in ~30 percent of tumors. LKB1 suppression or depletion leads to enhanced tumorigenesis, increased immunosuppression, and reduced response to immune checkpoint inhibitors. Despite the importance of LKB1 negative regulation on tumor growth and aggressiveness, there is no known upstream regulator of LKB1. Through a rationally designed drug screen, we identified tankyrases (TNKS; TNKS1, or PARP5A, and TNKS2, or PARP5B), as upstream negative regulators of LKB1. TNKS belong to the closely related members of the poly (ADP-ribose) polymerase (PARP) family that adds ADP-ribose moieties to target proteins using ?-NAD+ as substrate, termed PARsylation. PARsylation of target proteins by TNKS typically leads to ubiquination and proteasome degradation, such as AXIN (which sequesters APC/?-catenin in Wnt signaling), PTEN, and telomerase. However, PARsylation of LKB1 does not lead to LKB1 degradation, but inhibit LKB1 activity. Overexpression of TNKS stimulates tumor cellular proliferation in vitro and enhances tumorigenesis in vivo that is LKB1-dependent. Lung tumors (both adenocarcinoma and squamous cell carcinoma) that overexpress TNKS1 in LKB1 expressing tumors portended to poorer prognosis. Given the fact that LKB1 mutant lung tumors are immune suppressed and respond poorly to immune checkpoint inhibitors, we hypothesize that TNKS potentially could be the negative regulator of LKB1 that induce immune suppression in LKB1 wild type tumors. We will test our hypothesis by: (1) determining the role of TNKS on conferring immunosuppression in the tumor microenvironment through the use of syngeneic orthotopic models and Genetically-Engineered Mouse Models (GEMMs) to better understand if TNKS levels confers immunosuppressive state in tumors; and (2) utilizing syngeneic orthotopic tumor models to evaluate the ability of TNKS expression to generate resistance to immunotherapy and determine the effectiveness of TNKS inhibition on enhancing immunotherapy response. Upon completion of this project, we will have a clearer understanding of the immunosuppressive state that is conferred by TNKS through the negative regulation of LKB1. This research will implicate TNKS as a target for clinical translation to improve current therapies in lung cancer.